AJP - Gastrointestinal and Liver Physiology, Vol 259, Issue 3 410-G419, Copyright © 1990 by American Physiological Society
Phosphoinositide metabolism in intestinal brush borders: stimulation of IP3 formation by guanine nucleotides and Ca2+
A. B. Vaandrager, M. C. Ploemacher and H. R. De Jonge
Department of Biochemistry I, Erasmus University Rotterdam, The Netherlands.
The potential role of polyphosphoinositide (PPI) metabolism as a
signal-transduction mechanism in apical membranes of polarized epithelial
cells was evaluated by examining the formation and breakdown of PPI in rat
intestinal brush-border membranes (BBM) prelabeled by intraluminal
injection of [3H]inositol in vivo or by [gamma-32P]ATP in vitro. Freshly
isolated BBM prelabeled with [3H]inositol contained higher amounts of
[3H]phosphatidylinositol 4,5-diphosphate compared with a basolateral
membrane (BLM) preparation (approximately 14 and 6.8% of total [3H]PPIs,
respectively) and were enriched in inositol lipid kinases, diacylglycerol
(DAG) kinase, and phosphomonoesterases degrading PPI, inositol
bisphosphate, and inositol triphosphate (IP3). In the presence of
nonhydrolysable GTP analogues and low Ca2+ (pCa 6-8) or at high Ca2+ alone
(pCa 4) endogenous pools of PPI were rapidly depleted by an intrinsic
PPI-specific phospholipase C apparently coupled to a GTP-binding protein (G
protein). Surprisingly, despite the assignment of most G protein-coupled
hormone receptors to the BLM, the capacity of isolated BBM to release
[3H]IP3 in response to Ca2+ or GTP gamma S appeared comparable to that in a
BLM preparation. Intestinal secretagogues acting through apical membrane
receptors (adenosine, heat-stable Escherichia coli toxin), however, were
unable to promote [3H]IP3 release in isolated BBM in the presence of GTP.
PPI metabolism in BBM may be coupled to receptors for as yet unidentified
secretagogues or may serve as an amplification mechanism for
hormone-stimulated PPI breakdown in BLM. The local release of DAG and IP3
at the interior of the intestinal microvilli likely plays a role in the
regulation of ion transport systems in microvillar membranes.
